Two expression vectors for the phage-displayed chicken monoclonal antibody

• Published in: Journal of immunological methods Vol. 280; no. 1; pp. 157 - 164
• Main Authors: Nakamura, Naoto, Shimokawa, Mariko, Miyamoto, Kazuyoshi, Hojyo, Shintaro, Horiuchi, Hiroyuki, Furusawa, Shuichi, Matsuda, Haruo
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 01.09.2003; Elsevier
• Subjects: Animals; Antibodies, immunoglobulins; Antibodies, Monoclonal - genetics; Antibodies, Monoclonal - isolation & purification; Antibody Specificity; Base Sequence; Biological and medical sciences; Cell fusion; Chicken monoclonal antibodies; Chickens - genetics; Chickens - immunology; DNA, Complementary - genetics; Expression vectors; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Genetic Vectors; Hybridomas - immunology; Immunoglobulin Variable Region - genetics; Immunoglobulin Variable Region - isolation & purification; Mice; Molecular immunology; Monoclonal antibodies; Peptide Library; Phage display; Prions - immunology; Recombinant Proteins - genetics; Recombinant Proteins - immunology; Recombinant Proteins - isolation & purification; Vλ; Chicken monoclonal antibodies; CHSF; CLSB; Cell fusion; mAb; PrP; Phage display; VH; CRB; scFv; CHB; CLF; Expression vectors; Vertebrata; Chicken; Monoclonal antibody; Aves; Veterinary; Gene expression; Vector
• ISSN: 0022-1759; 1872-7905
• DOI: 10.1016/S0022-1759(03)00204-7

We previously reported the development of chicken monoclonal antibodies (mAb) against mammalian-conserved molecules by cell fusion and phage display using the mouse mAb expression vector pPDS. However, chicken hybridomas produce relatively small amounts of antibody when compared with mouse hybridomas, and application of the pPDS may be limited in two-antibody assays with a mouse mAb because it contains mouse Cκ as a detection tag. To circumvent the above problems, two expression vectors were established and used to produce a functional recombinant chicken mAb. These vectors, which were designed to accommodate a single chain fragment of the variable region (scFv) of the antibody, contained a chicken Cλ and FLAG with or without 6× histidine sequences in the 3′ terminus of the scFv to serve as detection and purification tags. In this study, a prion protein (PrP)-specific chicken mAb (HUC2-13) was expressed as phage-displayed and soluble scFv mAb forms by using these vectors. The scFv mAbs expressed by these vectors exhibited the same antigen-binding specificity to PrP as that of the original HUC2-13, could be purified with ease, and used in combination with a mouse mAb. These results indicate that the methods described herein offer an alternative to chicken mAb production from hybridomas and immunized chicken splenocytes, and may contribute to the use of chicken mAb reagents in numerous fields.